The discovery of statins.

This year, the Lasker Foundation confers its Clinical Medical Research Award on Akira Endo for his isolation from fungi of statins, potent inhibitors of cholesterol synthesis in the liver. The introduction of statins to clinical practice has markedly reduced morbidity and mortality from atherosclerotic cardiovascular disease.

Low Admission Plasma Gelsolin Concentrations Identify Community-acquired Pneumonia Patients at High Risk for Severe Outcomes.

BACKGROUND: Plasma gelsolin (pGSN) is an abundant circulating protein that neutralizes actin exposed by damaged cells, modulates inflammatory responses, and enhances alveolar macrophage antimicrobial activity. We investigated whether adults with low pGSN at hospital admission for community-acquired pneumonia (CAP) were at high risk for severe outcomes. METHODS: Admission pGSN concentrations in 455 adults hospitalized with CAP were measured using enzyme-linked immunosorbent assay. Patients were grouped into the following 4 hierarchical, mutually exclusive categories based on maximum clinical severity experienced during their hospitalization: general floor care without intensive care unit (ICU) admission, invasive respiratory or vasopressor support (IRVS), or death; ICU care without IRVS or death; IRVS without death; or death. Admission pGSN concentrations were compared across these discrete outcome categories. Additionally, outcomes among patients in the lowest quartile of pGSN concentration were compared to those in the upper 3 quartiles. RESULTS: Overall, median (interquartile range) pGSN concentration was 38.1 (32.1, 45.7) µg/mL. Patients with more severe outcomes had lower pGSN concentrations (P = .0001); median values were 40.3 µg/mL for floor patients, 36.7 µg/mL for ICU patients, 36.5 µg/mL for patients receiving IRVS, and 25.7 µg/mL for patients who died. Compared to patients with higher pGSN concentrations, patients in the lowest quartile (pGSN ≤ 32.1 µg/mL) more often required IRVS (21.2% vs 11.7%, P = .0114) and died (8.8% vs 0.9%, P < .0001). CONCLUSIONS: Among adults hospitalized with CAP, lower pGSN concentrations were associated with more severe clinical outcomes. Future studies are planned to investigate possible therapeutic benefits of recombinant human pGSN in this population.

Filamin A interaction with the CXCR4 third intracellular loop regulates endocytosis and signaling of WT and WHIM-like receptors.

Warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome is a rare congenital immunodeficiency often caused by mutations in the last 10 to 19 C-terminal amino acids of CXCR4. These mutations impair CXCR4 internalization and increase responsiveness to CXCL12. The CXCR4 C-terminal domain (C-tail) also has a binding site for the actin-binding protein filamin A (FLNA); it is not known whether FLNA binds to WHIM CXCR4 mutants or whether this interaction is implicated in the hyperfunction of these receptors. Here we show that, in addition to interacting with the CXCR4 C-tail, FLNA interacted with a region in the receptor third intracellular loop (ICL3) spanning amino acids 238 to 246. This interaction involved specific FLNA repeats and was sensitive to Rho kinase inhibition. Deletion of the 238-246 motif accelerated CXCL12-induced wild-type (WT) receptor endocytosis but enabled CXCL12-mediated endocytosis and normalized signaling by the WHIM-associated receptor CXCR4(R334X). CXCL12 stimulation triggered CXCR4(R334X) internalization in FLNA-deficient M2 cells but not in the FLNA-expressing M2 subclone A7; this suggests a role for FLNA in stabilization of WHIM-like CXCR4 at the cell surface. FLNA increased ß-arrestin2 binding to CXCR4(R334X) in vivo, which provides a molecular basis for FLNA-mediated hyperactivation of WHIM receptor signaling. We propose that FLNA interaction with ICL3 is central for endocytosis and signaling of WT and WHIM-like CXCR4 receptors.

An adventitious interaction of filamin A with RhoGDI2(Tyr153Glu).

Filamin A (FLNA) is an actin filament crosslinking protein with multiple intracellular binding partners. Mechanical force exposes cryptic FLNA binding sites for some of these ligands. To identify new force-dependent binding interactions, we used a fusion construct composed of two FLNA domains, one of which was previously identified as containing a force-dependent binding site as a bait in a yeast two-hybrid system and identified the Rho dissociation inhibitor 2 (RhoGDI2) as a potential interacting partner. A RhoGDI2 truncate with 81 N-terminal amino acid residues and a phosphomimetic mutant, RhoGDI(Tyr153Glu) interacted with the FLNA construct. However, neither wild-type or full-length RhoGDI2 phosphorylated at Y153 interacted with FLNA. Our interpretation of these contradictions is that truncation and/or mutation of RhoGDI2 perturbs its conformation to expose a site that adventitiously binds FLNA and is not a bona-fide interaction. Therefore, previous studies reporting that a RhoGDI(Y153E) mutant suppresses the metastasis of human bladder cancer cells must be reinvestigated in light of artificial interaction of this point mutant with FLNA.

Plasma gelsolin improves lung host defense against pneumonia by enhancing macrophage NOS3 function.

Plasma gelsolin (pGSN) functions as part of the "extracellular actin-scavenging system," but its potential to improve host defense against infection has not been studied. In a mouse model of primary pneumococcal pneumonia, recombinant human pGSN (rhu-pGSN) caused enhanced bacterial clearance, reduced acute inflammation, and improved survival. In vitro, rhu-pGSN rapidly improved lung macrophage uptake and killing of bacteria (Streptococcus pneumoniae, Escherichia coli, and Francisella tularensis). pGSN triggers activating phosphorylation (Ser(1177)) of macrophage nitric oxide synthase type III (NOS3), an enzyme with important bactericidal functions in lung macrophages. rhu-pGSN failed to enhance bacterial killing by NOS3(-/-) macrophages in vitro or bacterial clearance in NOS3(-/-) mice in vivo. Prophylaxis with immunomodulators may be especially relevant for patients at risk for secondary bacterial pneumonia, e.g., after influenza. Treatment of mice with pGSN challenged with pneumococci on postinfluenza day 7 (the peak of enhanced susceptibility to secondary infection) caused a ∼15-fold improvement in bacterial clearance, reduced acute neutrophilic inflammation, and markedly improved survival, even without antibiotic therapy. pGSN is a potential immunomodulator for improving lung host defense against primary and secondary bacterial pneumonia.

From the bench to the field in low-cost diagnostics: two case studies.

Despite the growth of research in universities on point-of-care (POC) diagnostics for global health, most devices never leave the laboratory. The processes that move diagnostic technology from the laboratory to the field--the processes intended to evaluate operation and performance under realistic conditions--are more complicated than they might seem. Two case studies illustrate this process: the development of a paper-based device to measure liver function, and the development of a device to identify sickle cell disease based on aqueous multiphase systems (AMPS) and differences in the densities of normal and sickled cells. Details of developing these devices provide strategies for forming partnerships, prototyping devices, designing studies, and evaluating POC diagnostics. Technical and procedural lessons drawn from these experiences may be useful to those designing diagnostic tests for developing countries, and more generally, technologies for use in resource-limited environments.

FilGAP, a Rho- and ROCK-regulated GAP for Rac binds filamin A to control actin remodelling.

FilGAP is a newly recognized filamin A (FLNa)-binding RhoGTPase-activating protein. The GTPase-activating protein (GAP) activity of FilGAP is specific for Rac and FLNa binding targets FilGAP to sites of membrane protrusion, where it antagonizes Rac in vivo. Dominant-negative FilGAP constructs lacking GAP activity or knockdown of endogenous FilGAP by small interference RNA (siRNA) induce spontaneous lamellae formation and stimulate cell spreading on fibronectin. Knockdown of endogenous FilGAP abrogates ROCK-dependent suppression of lamellae. Conversely, forced expression of FilGAP induces numerous blebs around the cell periphery and a ROCK-specific inhibitor suppresses bleb formation. ROCK phosphorylates FilGAP, and this phosphorylation stimulates its RacGAP activity and is a requirement for FilGAP-mediated bleb formation. FilGAP is, therefore, a mediator of the well-established antagonism of Rac by RhoA that suppresses leading edge protrusion and promotes cell retraction to achieve cellular polarity.

An active biopolymer network controlled by molecular motors.

We describe an active polymer network in which processive molecular motors control network elasticity. This system consists of actin filaments cross-linked by filamin A (FLNa) and contracted by bipolar filaments of muscle myosin II. The myosin motors stiffen the network by more than two orders of magnitude by pulling on actin filaments anchored in the network by FLNa cross-links, thereby generating internal stress. The stiffening response closely mimics the effects of external stress applied by mechanical shear. Both internal and external stresses can drive the network into a highly nonlinear, stiffened regime. The active stress reaches values that are equivalent to an external stress of 14 Pa, consistent with a 1-pN force per myosin head. This active network mimics many mechanical properties of cells and suggests that adherent cells exert mechanical control by operating in a nonlinear regime where cell stiffness is sensitive to changes in motor activity. This design principle may be applicable to engineering novel biologically inspired, active materials that adjust their own stiffness by internal catalytic control.

Cystic fibrosis transmembrane conductance regulator interacts with multiple immunoglobulin domains of filamin A.

Mutations of the chloride channel cystic fibrosis transmembrane conductance regulator (CFTR) that impair its apical localization and function cause cystic fibrosis. A previous report has shown that filamin A (FLNa), an actin-cross-linking and -scaffolding protein, interacts directly with the cytoplasmic N terminus of CFTR and that this interaction is necessary for stability and confinement of the channel to apical membranes. Here, we report that the CFTR N terminus has sequence similarity to known FLNa-binding partner-binding sites. FLNa has 24 Ig (IgFLNa) repeats, and a CFTR peptide pulled down repeats 9, 12, 17, 19, 21, and 23, which share sequence similarity yet differ from the other FLNa Ig domains. Using known structures of IgFLNa.partner complexes as templates, we generated in silico models of IgFLNa.CFTR peptide complexes. Point and deletion mutants of IgFLNa and CFTR informed by the models, including disease-causing mutations L15P and W19C, disrupted the binding interaction. The model predicted that a P5L CFTR mutation should not affect binding, but a synthetic P5L mutant peptide had reduced solubility, suggesting a different disease-causing mechanism. Taken together with the fact that FLNa dimers are elongated ( approximately 160 nm) strands, whereas CFTR is compact (6 approximately 8 nm), we propose that a single FLNa molecule can scaffold multiple CFTR partners. Unlike previously defined dimeric FLNa.partner complexes, the FLNa-monomeric CFTR interaction is relatively weak, presumptively facilitating dynamic clustering of CFTR at cell membranes. Finally, we show that deletion of all CFTR interacting domains from FLNa suppresses the surface expression of CFTR on baby hamster kidney cells.

Filamin is essential in actin cytoskeletal assembly mediated by p21-activated kinase 1.

The serine/threonine kinase p21-activated kinase 1 (Pak1) controls the actin cytoskeletal and ruffle formation through mechanisms that are independent of GTPase activity. Here we identify filamin FLNa as a Pak1-interacting protein through a yeast two-hybrid screen using the amino terminus of Pak1 as a bait. FLNa is stimulated by physiological signalling molecules to undergo phosphorylation by Pak1 and to interact and colocalize with endogenous Pak1 in membrane ruffles. The ruffle-forming activity of Pak1 is functional in FLNa-expressing cells but not in FLNa-deficient cells. In FLNa, the Pak1-binding site involves tandem repeat 23 in the carboxyl terminus and phosphorylation takes place on serine 2152. The FLNa-binding site in Pak1 is localized between amino acids 52 and 132 in the conserved Cdc42/Rac-interacting (CRIB) domain; accordingly, FLNa binding to the CRIB domain stimulates Pak1 kinase activity. Our results indicate that FLNa may be essential for Pak1-induced cytoskeletal reorganization and that the two-way regulatory interaction between Pak1 and FLNa may contribute to the local stimulation of Pak1 activity and its targets in cytoskeletal structures.

Low Plasma Gelsolin Concentrations in Chronic Granulomatous Disease.

Plasma gelsolin (pGSN) is the secreted isoform of an intracellular actin remodeling protein found in high concentrations in human plasma. Clinical studies demonstrate reduced pGSN concentrations in several disease states, including severe trauma, burns, and sepsis. Markedly decreased pGSN concentrations in these conditions precede and predict adverse clinical outcomes. In this study, we measured pGSN in patients with chronic granulomatous disease (CGD), a primary immunodeficiency characterized by recurrent infections and dysregulated inflammation. pGSN was quantified using a sandwich ELISA in plasma from healthy volunteers, clinically stable CGD patients, and X-linked CGD carriers and in sera from 12 CGD patients undergoing bone marrow transplantation. pGSN was also quantified in healthy volunteers challenged with intravenous endotoxin. pGSN concentrations were lower in CGD patients without active infection or systemic inflammation compared with healthy control subjects. In CGD patients undergoing bone marrow transplantation, pGSN concentrations increased significantly following successful transplant. X-linked carriers of CGD had normal pGSN. Despite reduction of pGSN in CGD patients, we did not detect significant changes in pGSN over 24 h following challenge of healthy volunteers with intravenous endotoxin (4 ng/kg) that elicited a febrile response. We describe, for the first time, significantly lower pGSN in clinically stable patients with CGD compared with age- and sex-matched healthy volunteers. Low pGSN levels in CGD patients significantly increased following bone marrow transplantation. X-linked carriers of CGD had normal pGSN. In healthy volunteers challenged with intravenous endotoxin, pGSN is not an acute phase reactant.

Plasma gelsolin and circulating actin correlate with hemodialysis mortality.

Plasma gelsolin (pGSN) binds actin and bioactive mediators to localize inflammation. Low pGSN correlates with adverse outcomes in acute injury, whereas administration of recombinant pGSN reduces mortality in experimental sepsis. We found that mean pGSN levels of 150 patients randomly selected from 10,044 starting chronic hemodialysis were 140 +/- 42 mg/L, 30 to 50% lower than levels reported for healthy individuals. In a larger sample, we performed a case-control analysis to evaluate the relationship of pGSN and circulating actin with mortality; pGSN levels were significantly lower in 114 patients who died within 1 yr of dialysis initiation than in 109 survivors (117 +/- 38 mg/L versus 147 +/- 42 mg/L, P < 0.001). pGSN levels had a graded, inverse relationship with 1-yr mortality, such that patients with pGSN < 130 mg/L experienced a > 3-fold risk for mortality compared with those with pGSN > or = 150 mg/L. The 69% of patients with detectable circulating actin had lower pGSN levels than those without (127 +/- 45 mg/L versus 141 +/- 36 mg/L, P = 0.026). Compared with patients who had elevated pGSN and no detectable actin, those with low pGSN levels and detectable actin had markedly increased mortality (odds ratio 9.8, 95% confidence interval 2.9 to 33.5). Worsening renal function correlated with pGSN decline in 53 subjects with CKD not on dialysis. In summary, low pGSN and detectable circulating actin identify chronic hemodialysis patients at highest risk for 1-yr mortality.

Recombinant Human Plasma Gelsolin Improves Survival and Attenuates Lung Injury in a Murine Model of Multidrug-Resistant Pseudomonas aeruginosa Pneumonia.

BACKGROUND: Plasma gelsolin (pGSN) is an abundant circulating protein quickly consumed by extensive tissue damage. Marked depletion is associated with later poor outcomes in diverse clinical circumstances. Repletion with recombinant human (rhu)-pGSN in animal models of inflammation lessens mortality and morbidity. METHODS: Neutropenic mice were treated with different meropenem doses ±12 mg of rhu-pGSN commencing 1 day before an intratracheal challenge with multidrug-resistant Pseudomonas aeruginosa. Survival, bacterial counts, and pulmonary pathology were compared between corresponding meropenem groups with and without rhu-pGSN. RESULTS: Overall survival was 35/64 (55%) and 46/64 (72%) in mice given meropenem without and with rhu-pGSN, respectively (Δ = 17%; 95% CI, 1-34). In control mice receiving meropenem 1250 mg/kg/d where the majority died, the addition of rhu-pGSN increased survival from 5/16 (31%) to 12/16 (75%) (Δ = 44%; 95% CI, 13-75). Survival with minor lung injury was found in 26/64 (41%) mice receiving only meropenem, vs 38/64 (59%) in mice given meropenem plus rhu-pGSN (Δ = 19%; 95% CI, 2-36). CONCLUSIONS: In a series of dose-ranging experiments, both mortality and lung injury were reduced by the addition of rhu-pGSN to meropenem against carbapenem-resistant P. aeruginosa. Rhu-pGSN offers a novel candidate therapy for antibiotic-resistant pneumonia.

Filling gaps in signaling to actin cytoskeletal remodeling.

A recent publication in the April 4 issue of Cell advances our understanding of stimulus response coupling leading to actin remodeling. It describes the identification of a novel membrane component Mig-2 that engages filamin A through a new intermediary, migfilin, to stimulate actin assembly and cell spreading on a substrate of extracellular matrix.

Blasts from the past.

With this issue of the JCI, we celebrate the 80th anniversary of the Journal. While 80 years is not a century, we still feel it is important to honor what the JCI has meant to the biomedical research community for 8 decades. To illustrate why the JCI is the leading general-interest translational research journal edited by and for biomedical researchers, we have asked former JCI editors-in-chief to reflect on some of the major scientific advances reported in the pages of the Journal during their tenures.

Ribosomal S6 kinase (RSK) regulates phosphorylation of filamin A on an important regulatory site.

The Ras-mitogen-activated protein (Ras-MAP) kinase pathway regulates various cellular processes, including gene expression, cell proliferation, and survival. Ribosomal S6 kinase (RSK), a key player in this pathway, modulates the activities of several cytoplasmic and nuclear proteins via phosphorylation. Here we report the characterization of the cytoskeletal protein filamin A (FLNa) as a membrane-associated RSK target. We show that the N-terminal kinase domain of RSK phosphorylates FLNa on Ser(2152) in response to mitogens. Inhibition of MAP kinase signaling with UO126 or mutation of Ser(2152) to Ala on FLNa prevents epidermal growth factor (EGF)-stimulated phosphorylation of FLNa in vivo. Furthermore, phosphorylation of FLNa on Ser(2152) is significantly enhanced by the expression of wild-type RSK and antagonized by kinase-inactive RSK or specific reduction of endogenous RSK. Strikingly, EGF-induced, FLNa-dependent migration of human melanoma cells is significantly reduced by UO126 treatment. Together, these data provide substantial evidence that RSK phosphorylates FLNa on Ser(2152) in vivo. Given that phosphorylation of FLNa on Ser(2152) is required for Pak1-mediated membrane ruffling, our results suggest a novel role for RSK in the regulation of the actin cytoskeleton.

Overregulation of conflicts hinders medical progress.

The revolution in medicine and technology over the past few decades is largely the result of partnerships--or a "harmony of interests"--between private companies and entrepreneurial scientists and clinicians. Regulations to prevent conflicts of interest by restricting medical education, medical research, expert advisory functions, or researcher ownership of inventions may have the unintended consequence of slowing medical progress.

A guide to the anti-innovation "pharmaphobia" narrative for the aspiring innovator.

Thomas P. Stossel is an American Cancer Society Professor of Medicine Emeritus at Harvard Medical School, Honorary Physician in the Hematology Division of Brigham & Women's Hospital, founding scientist of BioAegis Therapeutics, Visiting Scholar of The American Enterprise Institute and cofounder of Options for Children in Zambia.

Pediatric Dentists' Knowledge Concerning the Physician Payments Sunshine Act and Their Predictions of its Effects on Their Interactions with the Industry and its Impact on Patient Care.

PURPOSE: This study surveyed pediatric dentists' knowledge of the Physician Payments Sunshine Act (PPSA) and their impressions of its effect on their future interactions with the dental products industry and on their clinical practice. METHODS: Seven hundred seventy four practicing dentists responded to a survey, with 13 responding to a follow-up survey. RESULTS: Most respondents were unfamiliar (43 percent) or only vaguely familiar (33 percent) with the PPSA. In response to the required PPSA disclosures, 62 percent said they would see company representatives less often (37 percent indicated no change); 50 percent said they would attend fewer industry-sponsored continuing education events (49 percent indicated no change); and 57 percent indicated they would attend fewer promotional speaking events (43 percent indicated no change). Respondents indicated that self-banning detailing (68 percent), samples (72 percent), industry-sponsored continuing education events (81 percent), and promotional speaking (82 percent) would negatively impact the quality of care they are able to provide their patients. CONCLUSION: Pediatric dentists anticipate reducing or severing their existing ties to pharmaceutical, biotechnology, and device manufacturers in response to the Physician Payments Sunshine Act, despite believing that interactions with the dental products' industry enhance the quality of patient care they are able to provide their patients.